The aim of our research is to study regulation mechanisms of oncofetal gene expression in hepatocellular carcinogenesis in the mouse liver to determine whether specific oncogene expression occurs in normal and transformed hepatocytes and also to determine whether there are similarities in the regulation and structure of oncofetal protein genes (AFP) and oncogenes in the activated and repressed state. Using DNA-RNA hybridization techniques we have shown that AFP appears in DAB-fed mouse livers after 3 weeks of feeding and in CCl4-treated animals within 24 hours after treatment. These data indicate that activation of the AFP gene in adult liver is associated with early phases of hepatocarcinogenesis. Furthermore, since AFP activation occurs in liver regeneration, we propose that processes of AFP gene regulation occur in hepatocyte regeneration and during the early phases of hepatocarcinogenesis. However, upon reconstitution of the regenerating liver, the gene is again repressed as is indicated by reduced AFP mRNA levels. Our recent studies of oncogene expression have indicated that mRNA sequences complimentary to the C-Ha-ras-2 oncogene follow the same pattern of expression as the AFP gene in hepatocarcinogenesis and in CCl4-mediated regeneration. We will study the structure of these AFP genes and c-Ha-ras-2 oncogene at the DNA and chromatin level to determine if they have common features that could be the basis for similar regulatory mechanisms. Additional studies indicate that the repressed AFP gene undergoes structural changes during DAB and CCl4-mediated activation. We have found that the 5'-end flanking sequences of the AFP gene are more sensitive to DNase I digestion than specific regions within the structural gene at all ages, i.e., newborn, young adult and aging. We concluded that there are dynamic chromatin changes throughout the AFP gene during development, senescence and transformation of the hepatocyte. Studies are also in progress to determine whether gene activation (AFP and c-Ha-ras-2 sequences) involves changes in patterns of DNA methylation during hepatocarcinogenesis and liver regeneration.